Myelin-associated glycoprotein (MAG) is a member of the immunoglobulin gene superfamily that is localized in the periaxonal membranes of PNS and CNS myelin sheaths where it functions in glia-axon interactions and may be involved in transmitting signals from the axon to myelin forming cells. It occurs in two developmentally regulated isoforms with the large isoform, containing a longer cytoplasmic tail, predominating early in CNS development. The carbohydrate in MAG consists of a mixture of oligosaccharides, many of which are sialylated and sulfated, and which are currently being isolated and characterized. During this year, our studies on abnormalities of MAG expression in the dysmyelinating quaking mouse were completed. The results demonstrate a severe underexpression of the L-MAG isoform and several abnormalities of glycosylation, including increased sialylation, decreased content of the adhesion-related, HNK-1 epitope, and accumulation of a high mannose form of MAG. MAG expression in cultured oligodendrocytes and Schwann cells is being studied to determine factors that control its synthesis and posttranslational processing as well as investigating its function in intercellular adhesion and signaling. The immortalized Schwann cell lines generated in our laboratory express more MAG, MAG mRNA, P0 glycoprotein, P0 mRNA, sulfatide and galactocerebroside when their rate of growth is reduced by culturing in defined media or when the cells reach high density. The results suggest that these cells may be mimicking normal Schwann cells with regard to the necessity of down-regulating proliferation in order to differentiate toward myelination and should be useful for investigating factors controlling the expression of myelin constituents. Our previous findings, that GM3 ganglioside increases during normal oligodendrocyte differentiation in culture and that exogenous GM3 enhances the differentiation, were investigated further with regard to determining a possible effect of the ganglioside in modulating the activity of growth factor receptors. This year, we also examined an O-2A progenitor cell line (CG4) because of advantages that it may offer over primary cultures for investigating the functions of MAG and GM3. CG4 cells primarily express the L-MAG isoform and b-series gangliosides, and differ from primary oligodendrocytes by expressing very little GM3. The results suggest that the CG4 line will be useful for studying the functions of MAG and gangliosides in oligodendrocyte differentiation.